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I've been looking around the net looking for a nice protocol to validate micropipette calibrations using absorbance readings of a dye in solution. Does anyone have one they can share? I'd highly appreciate it!

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  • $\begingroup$ you mean absorbance? $\endgroup$
    – AliceD
    Jan 8, 2015 at 0:11
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    $\begingroup$ I just weigh water on an analytical balance and then compare the measured mass to what the mass should have been given the density of water. You should check the temperature of your water to be sure you have the right density. $\endgroup$
    – user137
    Jan 8, 2015 at 0:50
  • $\begingroup$ ^ I second this. I don't see why you would calibrate a micropipette using a spectro over gravimetric analysis. $\endgroup$
    – Macedon93
    Jan 8, 2015 at 1:03
  • $\begingroup$ I've never done it, but it seems to me like all you'd need to know is the concentration and extinction coefficient (at whatever wavelength) of the dye. $\endgroup$
    – canadianer
    Jan 8, 2015 at 2:10
  • $\begingroup$ Thanks for the feedback. We are just trying to come up with the fastest way possible of validating the accuracy of a robot that dispenses into 96 well plates within a given run as well as to check for variations from one run to another over the course of time. We are tight on time and do not have enough trained hands on deck for the gravinometric procedure. @Chris Stronks, I've edited accordingly. $\endgroup$
    – Yaritza
    Jan 8, 2015 at 3:53

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I see what you are getting at however this would be a very WRONG way of calibrating such an instrument. If dyes varied among trials it would throw off your calibration. Also, you would need to generate some sort of standard curve to equate absorbance to weight, since true calibration of these instruments is done by weighing the volumes they dispense (1g = 1ml water). The weight of water and the weight of your dye solution will not increase the same in terms of moles/g. Also a scale is far easier to calibrate that the detector in any spectrophotometer. Calibrate pipettes by weighing the volume of water they dispense. It should be done in a chamber that inhibits evaporation and you need a precision scale.

Here's a reliable guide for pipette calibration

enter link description here

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  • $\begingroup$ A regular 5 place balance with doors and some wet sponges on the edges should help minimize evaporation while you try to take measurements. $\endgroup$
    – virtualxtc
    Apr 26, 2018 at 19:37
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I found an article published by Beckman Coulter for measuring the accuracy of their liquid handling automation platforms. It could be used for hand-help pipettes as well

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  • $\begingroup$ Link-only answers are generally frowned upon on Stack Exchange sites. In time it is possible for links to atrophy (as your's has) and become unavailable, meaning that your answer is useless to users in the future. It would be best if you could provide the general details of your answer in your actual post, citing your link as a reference. $\endgroup$
    – virtualxtc
    Apr 26, 2018 at 19:35

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